Purification and Characterization of Mouse Liver Rhodanese

  • Lee, Chul-Young (Department of Biochemistry, College of Sciences, Hanyang University) ;
  • Hwang, Jae-Hoon (Department of Biochemistry, College of Sciences, Hanyang University) ;
  • Lee, Young-Seek (Department of Biochemistry, College of Sciences, Hanyang University) ;
  • Cho, Key-Seung (Department of Biochemistry, College of Sciences, Hanyang University)
  • 투고 : 1994.11.02
  • 발행 : 1995.03.31

초록

Rhodanese from mouse liver was purified to near homogeneity by ammonium sulfate precipitation, CM-Sephadex ion exchange, hydroxyapatite and Sephacryl S-200-HR gel filtration chromatographies with a purification of 776 folds. The molecular weight was determined by Sephadex G-150 gel filtration and found to be 34.8 KDa. SOS-PAGE showed molecular weight 34 KDa and two identical subunits splitting by aging for 3 weeks at $-70^{\circ}C$ the molecular weight of which was 17 KDa. The optimal pH of enzyme activity was 9.4 and the pI value of the enzyme was 6.6. Rhodanese showed the optimal reaction temperature of $25^{\circ}C$ and near linear increasing pattern until 10 min. incubation. $K_m$ values of rhodanese for KCN and $Na_{2}S_{2}O_{3}$ as substrates were 12.5 mM and 8.3 mM, respectively. Rhodanese activity was inhibited by more than 70% at a concentration of 100 ${\mu}M$ of $Ni^{2+}$, $Zn^{2+}$, $Cd^{2+}$, $Hg^{2+}$ and $Cu^{2+}$. Other metal ions, such as $Mn^{2+}$, $Mg^{2-}$, $Ca^{2+}$, and $Fe^{2+}$ showed no effect on rhodanese activity.

키워드

참고문헌

  1. Biochim. Biophys. Acta. v.956 Aird, B.A.;Horowitz, P.M. https://doi.org/10.1016/0167-4838(88)90294-4
  2. J. Biol. Chem. v.246 Blumenthal, K.M.;Heinrikson, R.L.
  3. Anal. Biochem. v.72 Bradford, M.
  4. J. Biol. Chem. v.245 Horowitz, P.M.;Detoma, F.
  5. Anal. Biochem. v.86 Horowitz, P.M. https://doi.org/10.1016/0003-2697(78)90803-5
  6. J. Biol. Chem. v.262 Horowitz, P.M.;Bowman, S.
  7. Biochem. Z. v.259 Lang, K.
  8. J. Biol. Chem. v.243 Leininger, K.R.;Westley, J.
  9. J. Biol. Chem. v.241 Mintel, R.;Westley, J.
  10. J. Protein Chem. v.5 Ogata, K.;Volini, M. https://doi.org/10.1007/BF01025422
  11. J. Biol. Chem. v.262 Ogata, K.;Dai, X.;Volini, M.
  12. Neurology v.37 Pallini, R.;Martelli, P.;Bardelli, A.M.;Guazzi, G.C.;Ferderico, A. https://doi.org/10.1212/WNL.37.12.1878
  13. Nature v.273 Ploegman, J.H.;Drent, G.;Kalk, K.H.;Hol, W.G.J.;Heinrikson, R.L.;Keim, P.S.;Weng, L.;Russell, J. https://doi.org/10.1038/273124a0
  14. J. Biol. Chem. v.253 Russell, J.;Weng, L.;Keim, P.S.;Heinrikson, R.L.
  15. Acta Chem. Scand. v.7 Sorbo, B.H. https://doi.org/10.3891/acta.chem.scand.07-1129
  16. Acta Chem. Scand. v.7 Sorbo, B.H. https://doi.org/10.3891/acta.chem.scand.07-1137
  17. Metabolic pathways, Vol.7 Sorbo, B.H.;Greenberg, D.M.(ed.)
  18. Patty's Industrial Hygiene and Toxicology, Vol. 2A Stokinger, H.E.;Clayton, G.D.(ed.);Clayton, F.E.(ed.)
  19. Annu. Rev. Biochem. v.41 Vallee, B.L.;Ulmer, D.D. https://doi.org/10.1146/annurev.bi.41.070172.000515
  20. J. Biol. Chem. v.242 Volini, M.;Detoma, F.;Westley, J.
  21. J. Biol. Chem. v.253 Volini, M.;Craven, D.;Ogata, K.
  22. J. Biol. Chem. v.243 Wang, S.F.;Violini, M.
  23. J. Biol. Chem. v.253 Weng, L.;Heinrikson, R.L.;Westley, J.
  24. J. Biol. Chem. v.243 Westley, J.;Green, J.R.
  25. J. Biol. Chem. v.243 Westley, J.
  26. Adv. Enzymol. v.39 Westley, J.